Hensen's node, the amniote equivalent of the amphibian "Spemann's organizer", is the most important region of the very early, gastrulating embryo. Not only does it generate the midline organs of the body (notochord, somites, gut, floor plate of the neural tube), but is also responsible for inducing and patterning the whole of the central nervous system. Classical studies have claimed, based on morphological criteria, that when Hensen's node is ablated, notochord and somites form normally. This project consists of new investigation of this problem, taking advantage of the many molecular markers now available, to reveal the conditions that specify this unique region of the embryo as "the organizer" and set its cells aside from the rest of the primitive streak. The study will help to define the organizer in both molecular and embryological terms and to identify the conditions that lead to the regulation of expression of several genes in this region. Finally, the results will reveal which properties of the organizer are under independent control at very early stages of embryonic development. Using transplantation experiments combined with in situ hybridization and retroviral vectors to cause misexpression of specific genes, the following specific questions will be addressed: Do the notochord and somites form after node ablation? Does the stump of the primitive streak express node- specific markers (e.g. goosecoid, sonic hedgehog) after node ablation? After ablation of the node, can the stump of the primitive streak acquire neural inducing ability and/or the ability to induce extra digits in a host limb bud? Is the regenerated node left/right asymmetric (like the normal node)? Is the ability to regenerate node properties restricted to a specific portion of the primitive streak? If any of the above properties cannot be acquired by the primitive streak, can it be made to regenerate these properties if made to express appropriate node marker genes?